Legal claims defining the scope of protection, as filed with the USPTO.
1. An optical memory device, comprising: a photochromic fluorescent protein moiety that is capable of being converted from a first fluorescent state to a second fluorescent state by irradiation with a writing wavelength in an optical memory device, wherein said first fluorescent state has a first excitation wavelength maximum, and wherein said photochromic fluorescent protein moiety is capable of being converted from said second fluorescent state to said first fluorescent state by irradiation with an erasing wavelength in an optical memory device.
2. The optical memory device of claim 1, wherein said first fluorescent state and said second fluorescent state are substantially stable at room temperature.
3. The optical memory device of claim 1, wherein said photochromic fluorescent protein moiety is an Aequorea-related photochromic fluorescent protein moiety.
4. The optical memory device of claim 1, wherein said second fluorescent state has a second excitation wavelength maximum, said second excitation wavelength maximum being longer than said first excitation wavelength maximum.
5. The optical memory device of claim 3, wherein excitation of said second fluorescent state at said second excitation maximum produces an intensity that is at least four times greater than said intensity of said first fluorescent state at said second excitation wavelength maximum.
6. The optical memory device of claim 4, wherein excitation of said second fluorescent state at said second excitation wavelength maximum produces an intensity that is at least eight times greater than said intensity of said first fluorescent state at said second excitation wavelength maximum.
7. The optical memory device of claim 3, wherein said second fluorescent state has a second excitation wavelength maximum, said second excitation wavelength maximum being shorter than said first excitation wavelength maximum.
8. The optical memory device of claim 3, wherein said photochromic fluorescent protein moiety includes an amino acid substitution of T203.
9. The optical memory device of claim 3, wherein said photochromic fluorescent protein moiety includes an amino acid substitution T203F, T203Y, or T203S.
10. The optical memory device of claim 3, wherein said photochromic fluorescent protein moiety includes amino acid substitutions S65G/S72A/T203F, S65G/S72A/T203Y, or T203S/S205T.
11. The optical memory device of claim 3, wherein said photochromic fluorescent protein moiety is a single polypeptide.
12. The optical memory device of claim 3, further comprising a medium including a plurality of said photochromic fluorescent protein moieties distributed throughout a medium.
13. The optical memory device of claim 12, wherein said medium is configured as a planar surface or a volume.
14. The optical memory device of claim 13, wherein said medium further includes polyacrylamide.
15. The optical memory device of claim 12, wherein each of said photochromic fluorescent protein moieties is individually addressable in said medium.
16. An optical memory device comprising: an Aequorea-related photochromic fluorescent protein moiety which is capable of being converted from a first fluorescent state to a second fluorescent state by irradiation with a writing wavelength in an optical memory device, wherein said photochromic fluorescent protein moiety is capable of being converted from said second fluorescent state to said first fluorescent state by irradiation with an erasing wavelength in an optical memory device, and wherein a medium including a plurality of said photochromic fluorescent protein moieties distributed throughout said medium, said medium being configured as a planar surface or a volume, further wherein said first fluorescent state has a first emission wavelength maximum, said first fluorescent state is capable of being excited by a reading wavelength in an optical memory device.
17. The optical memory device of claim 16, wherein said first fluorescent state and said second fluorescent state are substantially stable at room temperature.
18. The optical memory device of claim 16, wherein said second fluorescent state has a second excitation wavelength maximum, said second excitation wavelength maximum being longer than said first excitation wavelength maximum.
19. The optical memory device of claim 16, wherein excitation of said second fluorescent state at a second excitation wavelength maximum produces an intensity that is at least four times greater than said intensity of said first fluorescent state at said second excitation wavelength maximum.
20. The optical memory device of claim 16, wherein said second fluorescent state has a second excitation wavelength maximum, said second excitation wavelength maximum being shorter than said first excitation wavelength maximum.
21. The optical memory device of claim 16, wherein said photochromic fluorescent protein moiety includes an amino acid substitution of T203.
22. The optical memory device of claim 16, wherein said photochromic fluorescent protein moiety includes an amino acid substitution T203F, T203Y, or T203S.
23. The optical memory device of claim 16, wherein said photochromic fluorescent protein moiety includes amino acid substitutions S65G/S72A/T203F, S65G/S72A/T203Y, or T203S/S205T.
24. The optical memory device of claim 16, wherein said medium further includes polyacrylamide.
25. The optical memory device of claim 16, further comprising a medium including a plurality of said photochromic fluorescent protein moieties distributed throughout said medium.
26. The optical memory device of claim 24, wherein each of said photochromic fluorescent protein moieties is individually addressable in said optical memory device.
27. A composition of matter, comprising: a photochromic fluorescent protein moiety capable of being converted from a first fluorescent state to a second fluorescent state by irradiation with a writing wavelength in an optical memory device, wherein said first fluorescent state has a first excitation wavelength maximum, said second fluorescent state has a second excitation wavelength maximum, excitation of said second fluorescent state at said second wavelength excitation maximum produces an intensity that is at least four times greater than said intensity of said first fluorescent state at said second excitation wavelength maximum.
28. The composition of matter of claim 27, wherein said first fluorescent state and said second fluorescent state are substantially stable at room temperature.
29. The composition of matter of claim 27, wherein said photochromic fluorescent protein moiety is an Aequorea-related photochromic fluorescent protein moiety.
30. The composition of matter of claim 27, wherein said photochromic fluorescent protein moiety includes an amino acid substitution of T203.
31. The composition of matter of claim 27, wherein said photochromic fluorescent protein moiety includes an amino acid substitution T203F, T203Y, or T203S.
32. The composition of matter of claim 27, wherein said photochromic fluorescent protein moiety includes amino acid substitutions S65G/S72A/T203F, S65G/S72A/T203Y, or T203S/S205T.
33. The composition of matter of claim 27, wherein said photochromic fluorescent protein moiety is a single polypeptide.
34. The composition of matter of claim 27, wherein said photochromic fluorescent protein moiety is capable of being converted from said second fluorescent state to said first fluorescent state by irradiation with an erasing wavelength in an optical memory device.
35. The composition of matter of claim 27, wherein said second excitation wavelength maximum is longer than said first excitation wavelength maximum.
36. The composition of matter of claim 27, wherein said second fluorescent state has a second emission wavelength maximum and said first fluorescent state has a first emission wavelength maximum, wherein said second emission wavelength maximum is shorter than said first emission wavelength maximum.
37. The composition of matter of claim 27, wherein excitation of said second fluorescent state at said second excitation wavelength maximum produces an intensity that is at least 8 times greater than said intensity of said first fluorescent state at said second excitation wavelength maximum.
38. A method for storing and recovering information comprising: addressing a photochromic fluorescent protein moiety which is capable of being converted from a first fluorescent state to a second fluorescent state by irradiation with a writing wavelength in an optical memory device, wherein said first fluorescent state has a first excitation wavelength maximum, and wherein said photochromic fluorescent protein moiety is capable of being converted from said second fluorescent state to said first fluorescent state by irradiation with an erasing wavelength in an optical memory device, in a medium including a plurality of said photochromic fluorescent protein moieties; exposing said photochromic fluorescent protein moiety to said writing wavelength; irradiating said photochromic fluorescent protein moiety with a reading wavelength in an optical memory device; and detecting an output to determine whether said photochromic fluorescent protein moiety is in said first fluorescent state or second fluorescent state.
39. The method of claim 38 wherein, said first fluorescent state and said second fluorescent state are substantially stable at room temperature.
40. The method of claim 38, wherein said photochromic fluorescent protein moiety is an Aequorea-related photochromic fluorescent protein moiety.
41. The method of claim 40, wherein said second fluorescent state has a second excitation wavelength maximum, said second excitation wavelength maximum being longer than said first excitation wavelength maximum.
42. The optical memory device of claim 41, wherein excitation of said second fluorescent state at said second excitation wavelength maximum produces an intensity that is at least four times greater than said intensity of said first fluorescent state at second excitation wavelength maximum.
43. The method of claim 40, wherein said second fluorescent state has a second excitation wavelength maximum, said second excitation wavelength maximum being shorter than said first excitation wavelength maximum.
44. The method of claim 43, wherein detecting said output includes measuring an emission wavelength.
45. The method of claim 40, wherein said photochromic fluorescent protein moiety includes an amino acid substitution of T203.
46. The method of claim 40, wherein said photochromic fluorescent protein moiety includes an amino acid substitution T203F, T203Y, or T203S.
47. The method of claim 40, wherein said photochromic fluorescent protein moiety includes amino acid substitutions S65G/S72A/T203F, S65G/S72A/T203Y, or T203S/S205T.
48. The method of claim 38, wherein said photochromic fluorescent protein moiety is a single polypeptide.
49. The method of claim 38, wherein said medium is configured as a planar surface or a volume.
50. The method of claim 49, wherein said medium further includes polyacrylamide.
51. The method of claim 38, wherein each of said photochromic fluorescent protein moieties is individually addressable.
52. A method of improving a photochromic response of a photochromic fluorescent protein moiety, comprising: growing a plate of bacteria containing a large number of mutations that express a photochromic fluorescent protein moiety to provide a plurality of colonies; exposing said plurality of colonies to a first excitation wavelength and measuring an intensity of a resulting first emission I(A1) from said exposure to said first excitation wavelength of a colony; exposing said plurality of colonies to a second excitation wavelength and measuring an intensity of a resulting second emission I(B1) from said exposure to said second excitation wavelength of a colony; exposing said plurality of colonies to an isomerization wavelength; exposing said plurality of colonies to said first excitation wavelength and measuring an intensity of a first emission I(A2) from said exposure to said first excitation wavelength for a colony; exposing said plurality of colonies to said second excitation wavelength and measuring an intensity of a second emission I(B2) from said exposure to said second excitation wavelength for a colony; determining said ratio of emission intensities from said colony before and after exposure to said isomerization wavelength; and selecting said colony having improved photochromic response of said photochromic fluorescent protein moiety if said ratio of emission intensities is substantially different from an average ratio of emission intensities for said plurality of colonies.
53. The method of claim 52, wherein said photochromic fluorescent protein moiety is an Aequorea-related photochromic fluorescent protein moiety.
54. The method of claim 52, wherein said determining step includes calculating said ratio I(A2)/I(A1), l(B2)/I(B1), or I(A2)I(B1)/I(A1)I(B2).
55. The method of claim 52, further comprising exposing said plurality of colonies to an initial wavelength prior to said first exposing and measuring step.
56. The method of claim 52, wherein said exposing and measuring steps are performed with a digital imaging system.
57. The method of claim 52, further comprising picking up a portion of said colony having improved photochromic response of said photochromic fluorescent protein moiety.
58. The method of claim 57, wherein said picking up step is performed robotically.
59. An isolated nucleic acid sequence which encodes a photochromic fluorescent protein moiety capable of being converted from a first fluorescent state to a second fluorescent state by irradiation with a writing wavelength in an optical memory device, wherein said first fluorescent state has a first excitation wavelength maximum, said second fluorescent state has a second excitation wavelength maximum, excitation of said second fluorescent state at said second excitation wavelength maximum produces an intensity that is at least four times greater than said intensity of said first fluorescent state at said second excitation wavelength maximum.
60. The nucleic acid sequence of claim 59, wherein said first fluorescent state and said second fluorescent state are substantially stable at room temperature.
61. The nucleic acid sequence of claim 59, wherein said photochromic fluorescent protein moiety is an Aequorea-related photochromic fluorescent protein moiety.
62. The nucleic acid sequence of claim 61, wherein said photochromic fluorescent protein moiety is capable of being converted from said second fluorescent state to said first fluorescent state by irradiation with an erasing wavelength in an optical memory device.
63. The nucleic acid sequence of claim 61, said second excitation wavelength maximum is longer than said first excitation wavelength maximum.
64. The nucleic acid sequence of claim 61, said second excitation wavelength maximum is shorter than said first excitation wavelength maximum.
65. The nucleic acid sequence of claim 61, wherein said photochromic fluorescent protein moiety includes an amino acid substitution of T203.
66. The nucleic acid sequence of claim 61, wherein said photochromic fluorescent protein moiety includes an amino acid substitution T203F, T203Y, or T203S.
67. The nucleic acid sequence of claim 61, wherein said photochromic fluorescent protein moiety includes amino acid substitutions S65G/S72A/T203F, S65G/S72A/T203Y, or T203S/S205T.
68. An expression vector containing said nucleic acid sequence of claim 59.
69. An expression vector comprising expression control sequences operatively linked to a nucleic acid sequence coding for said expression of a photochromic fluorescent protein moiety capable of being converted from a first fluorescent state to a second fluorescent state by irradiation with a writing wavelength in an optical memory device, wherein said first fluorescent state has a first excitation wavelength maximum, said second fluorescent state has a second excitation wavelength maximum, excitation of said second fluorescent state at said second excitation wavelength maximum produces an intensity that is at least 4 times greater than said intensity of said first fluorescent state at said second excitation wavelength maximum.
70. The expression vector of claim 69, wherein said first fluorescent state and said second fluorescent state are substantially stable at room temperature.
71. The expression vector of claim 69, wherein said photochromic fluorescent protein moiety is an Aequorea-related photochromic fluorescent protein moiety.
72. The expression vector of claim 71, wherein said photochromic fluorescent protein moiety includes an amino acid substitution of T203.
73. The expression vector of claim 71, wherein said photochromic fluorescent protein moiety includes an amino acid substitution T203F, T203Y, or T203S.
74. The expression vector of claim 71, wherein said photochromic fluorescent protein moiety includes amino acid substitutions S65G/S72A/T203F, S65G/S72A/T203Y, or T203S/S205T.
75. The expression vector of claim 69, adapted for function in a prokaryotic cell.
76. The expression vector of claim 69, adapted for function in a eukaryotic cell.
77. A host cell transfected with an expression vector, comprising: an expression control sequence operatively linked to a sequence coding for said expression of a photochromic fluorescent protein moiety capable of being converted from a first fluorescent state to a second fluorescent state by irradiation with a writing wavelength in an optical memory device, wherein said first fluorescent state has a first excitation wavelength maximum, said second fluorescent state has a second excitation wavelength maximum, excitation of said second fluorescent state at said second excitation wavelength maximum produces an intensity that is at least four times greater than said intensity of said first fluorescent state at said second excitation wavelength maximum.
78. The host cell of claim 77, wherein said first fluorescent state and said second fluorescent state are substantially stable at room temperature.
79. The host cell of claim 77, wherein said photochromic fluorescent protein moiety is an Aequorea-related photochromic fluorescent protein moiety.
80. The host cell of claim 79, wherein said photochromic fluorescent protein moiety includes an amino acid substitution of T203.
81. The host cell of claim 79, wherein said photochromic fluorescent protein moiety includes an amino acid substitution T203F, T203Y, or T203S.
82. The host cell of claim 79, wherein said photochromic fluorescent protein moiety includes amino acid substitutions S65G/S72A/T203F, S65G/S72A/T203Y, or T203S/S205T.
83. The host cell of claim 77, wherein said cell is a prokaryote.
84. The host cell of claim 77, wherein said cell is E. coli.
85. The host cell of claim 77, wherein said cell is a eukaryotic cell.
86. The host cell of claim 85, wherein said cell is a yeast cell.
87. The host cell of claim 85, wherein said cell is a mammalian cell.
Complete technical specification and implementation details from the patent document.
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Unknown
April 4, 2000
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